The broad, long-term objective of this research proposal is to establish vascular smooth muscle cell (SMC)-derived carbon monoxide (CO) as a novel and biologically important messenger molecule that regulates vascular homeostasis at sites of vessel wall injury. We have recently discovered that SMC generate CO from the catabolism of heme oxygenase-1 (HO-1) and that specific humoral and hemodynamic factors encountered in the microenvironment of vascular injury induces HO-1 gene expression and CO release. Based on these findings, we propose to study the biological effects of CO on vascular SMC function. These studies will determine the effect of both exogenously administered and endogenously-derived CO on blood vessel tone, SMC proliferation, and SMC collagen synthesis. In all instances where CO is shown to alter SMC function we will determine the molecular basis of this effect. Since we have shown that Co stimulates soluble guanylate cyclase activity in vascular SMC, we will investigate whether cGMP and protein kinase G (PKG) and/or A (PKA) mediates the biological effects of CO. We will further explore the role of CO as an autocrine regulator of SMC function by generating SMC that over-express HO-1 enzyme. If HO-1 transfected SMC have altered biological responses we will attempt to restore responses by treating SMC with specific metalloprotoporphyrin HO inhibitors or by treating SMC with HO-1 antisense oligodeoxynucleotides. Finally, we will investigate the role of CO in regulating intimal thickening and vascular reactivity in the rat carotid artery balloon injury model. We will determine whether the local inhibition or over-expression of HO-1 activity and CO production regulates lesion formation and vascular tone at sites of vascular injury. In addition, we will determine whether cGMP and PKG/PKA are responsible for any of the in vivo effects of CO. It is anticipated that these studies will (a) establish CO as a novel SMC-derived signaling molecule that regulates SMC contraction, proliferation and collagen synthesis, and (b) lead to the development of novel therapeutic strategies in treating occlusive vascular disease.